Activated AMPK-mediated glucose uptake and mitochondrial dysfunction is critically involved in the glutamate-induced oxidative injury in HT22 cell.

BACKGROUND: Accumulation of glutamate damages neurons via the reactive oxygen species (ROS) injury, which was involved in the development of neurodegenerative diseases. However, the mechanism of neuronal oxidative stress damage caused by glutamate and the intervention targets still needs to be further studied. This study explored whether 5' adenosine monophosphate-activated protein kinase (AMPK)-induced glucose metabolic and mitochondrial dysfunction were related to glutamate-dependent ROS injury of the neuron. METHODS: Neuronal oxidative stress injury was induced by glutamate treatment in HT-22 cells. Western blotting was used to evaluate the phosphorylation of the AMPK. The XF24 Flux Analyzer was used to measure the effect of glutamate and Compound C (a well-known pharmacological inhibitor of AMPK phosphorylation) on the cellular oxygen consumption rate (OCR) of HT-22 cells. Glucose uptake, intracellular ROS, mitochondrial potential, apoptosis and cell viability were quantified using biochemical assays. RESULTS: Glutamate caused the phosphorylation of AMPK and subsequently promoted the glucose uptake. Furthermore, AMPK-mediated glucose uptake enhanced OCR and increased the intracellular ROS levels in neurons. The pharmacological inhibition of AMPK phosphorylation by Compound C attenuated glutamate-induced toxicity in HT22 cells by regulating the glucose uptake/mitochondrial respiration/ROS pathway. CONCLUSIONS: The AMPK phosphorylation/glucose uptake/mitochondrial respiration/ROS pathway was involved in glutamate-induced excitotoxic injury in HT22 cells. The inhibition of AMPK phosphorylation may be a potential target for the development of therapeutic agents for treating the glutamate-induced neurotoxicity.

Artemisinin improves neurocognitive deficits associated with sepsis by activating the AMPK axis in microglia.

Sepsis is life-threatening organ dysfunction due to dysregulated systemic inflammatory and immune response to infection, often leading to cognitive impairments. Growing evidence shows that artemisinin, an antimalarial drug, possesses potent anti-inflammatory and immunoregulatory activities. In this study we investigated whether artemisinin exerted protective effect against neurocognitive deficits associated with sepsis and explored the underlying mechanisms. Mice were injected with LPS (750 µg · kg-1 · d-1, ip, for 7 days) to establish an animal model of sepsis. Artemisinin (30 mg · kg-1 · d-1, ip) was administered starting 4 days prior LPS injection and lasting to the end of LPS injection. We showed that artemisinin administration significantly improved LPS-induced cognitive impairments assessed in Morris water maze and Y maze tests, attenuated neuronal damage and microglial activation in the hippocampus. In BV2 microglial cells treated with LPS (100 ng/mL), pre-application of artemisinin (40 µΜ) significantly reduced the production of proinflammatory cytokines (i.e., TNF-α, IL-6) and suppressed microglial migration. Furthermore, we revealed that artemisinin significantly suppressed the nuclear translocation of NF-κB and the expression of proinflammatory cytokines by activating the AMPKα1 pathway; knockdown of AMPKα1 markedly abolished the anti-inflammatory effects of artemisinin in BV2 microglial cells. In conclusion, atemisinin is a potential therapeutic agent for sepsis-associated neuroinflammation and cognitive impairment, and its effect is probably mediated by activation of the AMPKα1 signaling pathway in microglia.

Silencing of circFoxO3 Protects HT22 Cells from Glutamate-Induced Oxidative Injury via Regulating the Mitochondrial Apoptosis Pathway.

Recent studies demonstrated that FoxO3 circular RNA (circFoxO3) plays an important regulatory role in tumourigenesis and cardiomyopathy. However, the role of circFoxO3 in neurodegenerative diseases remains unknown. The aim of this study was to examine the possible role of circFoxO3 in neurodegenerative diseases and the underlying mechanisms. To model human neurodegenerative conditions, hippocampus-derived neurons were treated with glutamate. Using molecular and cellular biology approaches, we found that circFoxO3 expression was significantly higher in the glutamate treatment group than that in the control group. Furthermore, silencing of circFoxO3 protected HT22 cells from glutamate-induced oxidative injury through the inhibition of the mitochondrial apoptotic pathway. Collectively, our study demonstrates that endogenous circFoxO3 plays a key role in inducing apoptosis and neuronal cell death and may act as a novel therapeutic target for neurodegenerative diseases.

DNA methylation inhibitor, decitabine, promotes MGC803 gastric cancer cell migration and invasion via the upregulation of NEDD4­1.

Gastric cancer is the fourth most common cancer type and the second leading cause of cancer­associated mortality worldwide. Metastasis is a crucial feature of its progression. DNA methylation provides a key epigenetic signature in the epigenetic regulation pathway, and is implicated in transcriptional regulation. CpG sites, which are associated with gene transcriptional activity, are underrepresented in the mammalian genome and tend to be clustered within CpG islands (CGIs) located in the vicinity of the transcription start sites of the majority of the protein­coding genes in humans. The DNA methylation inhibitor, decitabine (DAC), has been demonstrated to be active in hematological disorders. The majority of previous studies in cancer cells demonstrated that DAC inhibits cell proliferation and the motility of the cells. However, since demethylation across the entire genome alters the expression of a large number of genes, the effects of DAC in different tumor cell types are difficult to accurately predict. Neural precursor cell­expressed, developmentally downregulated (NEDD)4­1, a member of the NEDD4 family, which belongs to the E3­ubiquitin ligase family, was reported to be highly expressed in a wide range of tumor types, and it activates the phosphoinositide 3­kinase/Akt pathway by degrading phosphatase and tensin homolog. NEDD4­1 promotes the migration and invasion of glioma cells via the ubiquitination and subsequent degradation of cyclic nucleotide­Ras guanine nucleotide exchange factors (CNrasGEFs). In gastric cardia adenocarcinoma, NEDD4­1 acts as an exceptional prognostic biomarker. In the present study, DAC was revealed to promote the invasive properties of MGC803 gastric cancer cells. NEDD4­1 targeted the CNrasGEF­mediated DAC invasion­promoting activity in MGC803 cells, and CGI methylation in neither the NEDD4 promoter nor the first intron was demonstrated to be associated with this effect. The results of the present study revealed that DAC exerts variable effects in different gastric cancer cell lines and may provide a reference for DAC administration in the clinic.

Calcium Ion Flow Permeates Cells through SOCs to Promote Cathode-Directed Galvanotaxis.

Sensing and responding to endogenous electrical fields are important abilities for cells engaged in processes such as embryogenesis, regeneration and wound healing. Many types of cultured cells have been induced to migrate directionally within electrical fields in vitro using a process known as galvanotaxis. The underlying mechanism by which cells sense electrical fields is unknown. In this study, we assembled a polydimethylsiloxane (PDMS) galvanotaxis system and found that mouse fibroblasts and human prostate cancer PC3 cells migrated to the cathode. By comparing the effects of a pulsed direct current, a constant direct current and an anion-exchange membrane on the directed migration of mouse fibroblasts, we found that these cells responded to the ionic flow in the electrical fields. Taken together, the observed effects of the calcium content of the medium, the function of the store-operated calcium channels (SOCs) and the intracellular calcium content on galvanotaxis indicated that calcium ionic flow from the anode to the cathode within the culture medium permeated the cells through SOCs at the drift velocity, promoting migration toward the cathode. The RTK-PI3K pathway was involved in this process, but the ROCK and MAPK pathways were not. PC3 cells and mouse fibroblasts utilized the same mechanism of galvanotaxis. Together, these results indicated that the signaling pathway responsible for cathode-directed cellular galvanotaxis involved calcium ionic flow from the anode to the cathode within the culture medium, which permeated the cells through SOCs, causing cytoskeletal reorganization via PI3K signaling.

OCT4B modulates OCT4A expression as ceRNA in tumor cells.

OCT4 is an essential transcription factor for maintaining the self-renewal and the pluripotency of embryonic stem cells (ESCs). The human OCT4 gene can generate three mRNA isoforms (OCT4A, OCT4B and OCT4B1) by alternative splicing. OCT4A protein is a transcription factor for the stemness of ESCs, while the function of OCT4B isoforms remains unclear. Most types of cancer express a relatively low level of OCT4 protein, particularly the OCT4B isoforms. In the present study, we found that OCT4A and OCT4B mRNA were co-expressed in several types of tumor cell lines and tumor samples, and we demonstrated that OCT4B functioned as a non-coding RNA, modulating OCT4A expression in an miRNA-dependent manner [competing endogenous RNA (ceRNA) regulation] at the post-transcription level in the tumor cell lines. This is the first time that ceRNA regulation was observed among spliced isoforms of one gene, and may pave the way for identification of new targets for cancer treatment.

MicroRNA-124 regulates TGF-α-induced epithelial-mesenchymal transition in human prostate cancer cells.

Transforming growth factor-α (TGF-α) is upregulated in advanced stages of prostate cancer and strongly correlated with metastasis. However, the effect of TGF-α on epithelial-mesenchymal transition (EMT) in prostate cancer and the underlying mechanisms remain unclear. Recently, microRNAs have emerged as new regulators of EMT. This study found that treatment of DU145 cells with TGF-α suppressed the expression of epithelial marker E-cadherin and increased the expression of mesenchymal marker Vimentin as well as changed the cell morphology from cobblestone shape to spindle shape. The level of miR-124 was downregulated by TGF-α in several different cancer cell lines. Enforced expression of miR-124 abolished TGF-α-induced EMT. Slug was proven to be a target of miR-124 and mediated the inhibitory effect of miR-124 on TGF-α-induced EMT. Furthermore, overexpression of miR-124 reduced the migratory and invasive capacity of TGF-α-treated DU145 cells. In conclusion, our findings suggest that miR-124 inhibits TGF-α-induced EMT in DU145 cells by targeting Slug. Thus, miR-124 may be a potential target for prostate cancer therapeutic intervention.

Gap junction permeability modulated by dopamine exerts effects on spatial and temporal correlation of retinal ganglion cells' firing activities.

Synchronized activities among retinal ganglion cells (RGCs) via gap junctions can be increased by exogenous dopamine (DA). During DA application, single neurons' firing activities become more synchronized with its adjacent neighbors. One intriguing question is how the enhanced spatial synchronization alters the temporal firing structure of single neurons. In the present study, firing activities of bullfrog's dimming detectors in response to binary pseudo-random checker-board flickering were recorded via a multi-channel recording system. DA was applied in the retina to modulate synchronized activities between RGCs, and the effect of DA on firing activities of single neurons was examined. It was found that, during application of DA, synchronized activities between single neuron and its neighboring neurons was enhanced. At the meantime, the temporal structures of single neuron spike train changed significantly, and the temporal correlation in single neuron's response decreased. The pharmacological study results indicated that the activation of D1 receptor might have effects on gap junction permeability between RGCs. Our results suggested that the dopaminergic pathway participated in the modulation of spatial and temporal correlation of RGCs' firing activities, and may exert critical effects on visual information processing in the retina.